1. Field of the Invention
The present invention relates to a dispenser to be used for the production of a DNA chip (DNA microarray) in which several thousands to not less than ten thousands kinds of different types of DNA fragments are aligned and fixed as minute spots at a high density on a base plate such as a microscopic glass slide, and a method for producing a DNA chip by using the dispenser.
2. Description of the Related Art
The method for analyzing the genetic structure has been remarkably progressed in recent years. A large number of genetic structures represented by those of the human genome have been clarified. The analysis of the genetic structure as described above uses a DNA chip (DNA microarray) in which several thousands to not less than ten thousands kinds of different types of DNA fragments are aligned and fixed as spots on a base plate such as a microscopic glass slide.
Those widely used as a method for forming the spots for the production of the DNA chip are based on a system such as the QUILL system, the pin and ring system, and the spring pin system in which a sample solution containing DNA fragments is supplied (stamped) onto the base plate by using a so-called pin. Even when any one of the foregoing methods is adopted, it is necessary to suppress the dispersion of the volume and the shape of each of the spots to be low so that the distance between the respective spots is maintained to be constant.
On the other hand, in order to realize a higher density, it is also greatly expected to develop a new method which is excellent in productivity and in which the shape control performance for the spot is satisfactory.
The QUILL system lies in a method in which a sample is stored in a recess formed at a pin tip, and the pin tip is allowed to make contact with the base plate so that the sample in the recess is transferred onto the base plate to form a minute spot. However, this system involves, for example, a problem of durability such that the pin tip is deformed or it is damaged by the contact with the base plate. Further, this system also involves, for example, a problem such that the sample stored in the recess is incompletely washed to facilitate the occurrence of cross-contamination.
The pin and ring system lies in a method in which a sample solution in a microplate is reserved with a ring, and then the sample in the ring is trapped by a pin tip so that the pin tip penetrates through the inside of the ring reserved with the solution to form a spot on the base plate. However, the sample, which can be reserved once, depends on the number of rings. Conventionally, the number of rings can not be increased. For this reason, in order to form several thousands to several tens of thousands of minute sample spots, it is also necessary to perform washing and drying steps several hundreds to several thousands of times. Therefore, it is difficult to say that the productivity is necessarily high.
The spring pin system lies in a method in which a sample adhered to a pin tip is transferred onto the base plate by pressing the pin tip against the base plate. A double pin structure containing a spring is used to mitigate the damage of the pin and the base plate while the sample is ejected. However, basically, only one time of spotting can be performed with one time of reserve. This system is inferior in productivity.
In all of the conventional methods for forming the minute spots, the sample solution is transported onto the base plate in a state of being exposed to the atmospheric air. Therefore, an inconvenience arises such that the sample is dried during the transport, and it is impossible to perform the spotting. A problem arises such that the extremely expensive sample solution is used with bad efficiency.
On the other hand, a method is also investigated by using the so-called ink-jet system which is practically used for printers. However, many tasks arise concerning the size and the cost when several thousands to several tens of thousands of individual flow passages corresponding to every sample are formed. Further, in the case of the ink-jet system, it is necessary to previously charge the sample in a pump before the spotting so that no bubble is formed. For this reason, a large amount of sample is required to effect the purge. Therefore, the efficiency of the use of the sample is extremely inferior. In general, in order to remove the bubble, it is preferable to move the liquid at a high speed in the flow passage including a pump chamber. As a result, the sample is agitated in the flow passage. When a delicate DNA solution is used as a sample, for example, DNA is damaged in some cases.
The present invention has been made taking the foregoing problems into consideration, an object of which is to provide a dispenser which comprises a large number of micropipettes arranged to makes it possible to form minute spots accurately at a high speed, which makes it possible to supply a solution to the respective micropipettes quickly and reliably, and which makes it possible to smoothly perform the steps from the supply of the solution to the supply onto a base plate.
Another object of the present invention is to provide a method for producing a DNA chip, which makes it possible to smoothly perform the steps from the supply of a solution to the supply onto a base plate and which makes it possible to improve the quality of the DNA chip and improve the yield.
According to the present invention, there is provided a dispenser comprising a plurality of arranged micropipettes each including a pouring port for pouring a sample solution from the outside, a cavity for pouring and charging the sample solution thereinto, and a discharge port for discharging the sample solution, formed on at least one or more substrates. The micropipette further includes a piezoelectric/electrostrictive element disposed on at least one wall surface of the substrate which forms the cavity so that the sample solution is movable in the cavity, and the sample solution is discharged from the discharge port of each of the micropipettes; wherein a pin, which protrudes upwardly, is provided at the pouring port of each of the micropipettes.
According to another aspect of the present invention, there is provided a method for producing a DNA chip, comprising the steps of using a dispenser comprising a plurality of arranged micropipettes each including a pouring port for pouring a sample solution from the outside, a cavity for pouring and charging the sample solution thereinto, and a discharge port for discharging the sample solution, formed on at least one or more substrates. The micropipette further includes a piezoelectric/electrostrictive element disposed on at least one wall surface of the substrate which forms the cavity so that the sample solution is movable in the cavity; and discharging the sample solution onto a base plate from the discharge port of each of the micropipettes to produce the DNA chip; wherein the dispenser to be used is provided with a pin protruding upwardly at the pouring port of each of the micropipettes.
Accordingly, a hole can be bored with the pin through a solution storage section of a cartridge positioned over the pouring port so that the solution stored in the solution storage section may be introduced into the pouring port.
That is, a cartridge, which is arranged with a large number of solution storage sections, is positioned over the dispenser, and the cartridge is moved toward the dispenser. At this time, a hole is bored with the pin through each of the solution storage sections. Accordingly, the solution stored in the solution storage section is introduced into the pouring port along the pin. By doing so, it is unnecessary to use any special apparatus when the sample solution is poured into the dispenser from the solution storage section of the cartridge. Thus, no sample solution remains in the special apparatus, and the efficiency of the use of the sample solution is not lowered.
In this process, it is also preferable that when the solution stored in the solution storage section is introduced into the pouring port, a gas is fed under pressure downwardly to each of the solution storage sections. Accordingly, it is possible to shorten the period of time required to pour the solution.
In the present invention, the solution stored in the solution storage section can be introduced into the pouring port by boring a hole through a film member coated to close the solution storage section of the cartridge positioned over the pouring port.
That is, a cartridge, which is arranged with a large number of solution storage sections, is coated with a film member to close the solution storage sections; the cartridge is positioned over the dispenser so that the film member is opposed to the dispenser; and the cartridge is moved toward the dispenser. During this process, a hole is bored with the pin through a portion of the film member corresponding to each of the solution storage sections. Accordingly, the solution stored in the solution storage section is introduced into the pouring port along the pin.
It is possible to carry out the process to bore the hole through the film member relatively conveniently, as compared with the process to bore the hole through the solution storage section of the cartridge. Therefore, it is easy to introduce the sample solution.
As described above, the dispenser according to the present invention can be used to supply the solution to the respective micropipettes quickly and reliably. It is possible to perform the steps from the supply of the solution to the supply onto the base plate.
The pin described above refers to a projection-shaped section having a portion protruding from the plane. It is preferable that the forward end is sharp. It is preferable that the arrangement position of each of the pouring ports of the micropipettes for constructing the dispenser is equal to the arrangement position of each of the solution storage sections of the cartridge provided with the solution storage sections. Alternatively, it is preferable that the arrangement pitch is an integral multiple of the arrangement pitch of the solution storage sections, or an integral fraction thereof.
The pin may be provided at a position included in the pouring port as viewed in plan view, or it may be provided at a circumferential edge of the pouring port. When the pin is provided at the position included in the pouring port as viewed in plan view, the hole, through which the sample solution is introduced, can be positioned just over the pouring port. Thus, it is possible to introduce the sample solution more reliably. When the pin is provided at the circumference of the pouring port, then it is easy to form the pin, and the production cost of the dispenser is reduced.
In the present invention, it is also preferable that a holding section for holding a pipette for pouring the solution from the pouring port or a tube for receiving the pipette is provided at a circumferential edge of the pouring port of each of the micropipettes for constructing the dispenser.
Accordingly, when the solution is poured into each of the micropipettes of the dispenser by using the pipette, the pipette or the tube for receiving the pipette is held by the holding section. Therefore, it is possible to pour the solution into the micropipette in a reliable manner. It is possible to effectively avoid, for example, any leakage of the solution.
Especially, at least the inner wall of the tube for receiving the pipette is subjected to a hydrophilic treatment. Thus, the solution, which is discharged from the pipette, can be reliably introduced into the pouring port of the micropipette without involving any bubble or the like.
In the present invention, it is also preferable that a scale for measuring an amount of liquid poured into the tube is formed at least at a part of the tube for receiving the pipette. It is also preferable that a portion provided with a projection and a portion provided with no projection are formed at positions of an identical distance from the pouring port on a part of an inner wall of the tube for receiving the pipette.
When the scale is formed, it is possible to measure and confirm the amount of the poured sample solution while pouring the sample solution, and the amount of the discharged sample solution while discharging the sample solution. Thus, the scale is useful to manage the production of the product and manage the quality. Further, the scale is effective to manage the liquid amount of a substitution solution or an intermediate solution in the case of the use of a method in which the substitution solution or the intermediate solution is previously charged, when the sample solution is poured and charged into the micropipette. As a result, it is possible to perform the substitution of the substitution solution certainly or the intermediate solution with the sample solution. Thus, it is possible to reduce the dispersion of the concentration of the sample solution to be supplied, and the quality of the product is improved.
When the portion provided with the projection and the portion provided with no projection are formed at the positions of the identical distance from the pouring port on the part of the inner wall of the tube for receiving the pipette, it is possible to perform the introducing operation while allowing the forward end of the pipette for introducing the sample solution to make contact with the projection. The injection position of the pipette can be always made to be constant, and the dispersion of the introducing operation is reduced.
The presence of the portion provided with the projection and the portion provided with no projection ensures the escape route for the gas during the pouring process. It is possible to perform the introducing operation without involving any bubble or the like. It is noted that such an effect is exhibited not only when the sample solution, the substitution solution or the like is introduced (poured). The foregoing arrangement is also effective when the pipetting is performed in order to remove an excessive amount of the sample solution, the substitution solution, or the intermediate solution.
In the present invention, it is preferable that a filter, which is formed with a large number of openings having an opening area of not more than an opening area of the discharge port, is attached between the pouring port and the tube for receiving the pipette, in order to remove any foreign matter in the sample solution to be poured. By doing so, it is possible to previously avoid the invasion of the foreign matter into the micropipette, the clogging of the discharge port or the like, and the failure of the supply of the sample solution.
In the present invention, it is also preferable that the dispenser further comprises a pitch-varying mechanism for varying an arrangement pitch of each of the micropipettes for constructing the dispenser.
Accordingly, the solution can be supplied to the dispenser while allowing the arrangement pitch of the respective micropipettes of the dispenser to conform to an arrangement pitch of respective pipettes of a solution supply means for supplying the solution to the dispenser; and the sample solution can be supplied from the dispenser onto the base plate while setting the arrangement pitch of the respective micropipettes of the dispenser to be a pitch which is different from the arrangement pitch of the respective pipettes of the solution supply means. It is possible to perform the steps from the supply of the solution to the supply onto the base plate continuously.
That is, in general, the supply (pouring or introduction) of the sample solution to the micropipette and the dispenser is often restricted by the size of the solution supply means or the cartridge having the solution storage section. It is inevitable to give a relatively large arrangement pitch for the respective pipettes or the pouring ports of the respective micropipette. On the other hand, when the sample solution is supplied onto the base plate, the small supply pitch is advantageous in many cases in view of the spot density and the number of spots capable of being supplied once. In such a situation, the dispenser according to the present invention is preferably adopted.
In the present invention, it is also preferable that a solution supply means, which is arranged with a large number of pipettes for supplying the solution to the dispenser and which has a pitch-varying mechanism for varying an arrangement pitch of the respective pipettes, is used; the solution is supplied to the solution supply means while allowing the arrangement pitch of the respective pipettes to conform to an arrangement pitch of solution storage sections of a cartridge; and the solution is supplied from the solution supply means to the dispenser while allowing the arrangement pitch of the respective pipettes to conform to an arrangement pitch of the micropipettes of the dispenser.
In this case, it is possible to smoothly perform the process for supplying, to the dispenser, the solution stored in the respective solution storage sections of the cartridge. It is possible to effectively shorten the period of time required for the production.
In the present invention, when the dispenser is used, it is also preferable to adopt the following procedure. That is, no pin is provided for the dispenser. A cartridge, which is arranged with a large number of solution storage sections, is positioned over the dispenser. A pin is externally used to bore a hole through each of the solution storage sections so that the solution stored in the solution storage section is introduced into the pouring port. In this case, a simple structure can be used for each of the micropipettes of the dispenser.
When the hydrophilic treatment is applied to the pouring port of the dispenser described above, the sample solution, which is supplied via the pouring port, can be smoothly introduced toward the cavity. Therefore, it is possible to shorten the period of time required to supply the sample solution.
The above and other objects, features, and advantages of the present invention will become more apparent from the following description when taken in conjunction with the accompanying drawings in which a preferred embodiment of the present invention is shown by way of illustrative example.